McIntyre, PJ, Collins, PM, Vrzal, L et al. (11 more authors) (2017) Characterization of three druggable hot-spots in the Aurora-A/TPX2 interaction using biochemical, biophysical and fragment-based approaches. ACS Chemical Biology, 12 (11). pp. 2906-2914. ISSN 1554-8929
Abstract
The mitotic kinase Aurora-A and its partner protein TPX2 (Targeting Protein for Xenopus kinesin-like protein 2) are overexpressed in cancers, and it has been proposed that they work together as an oncogenic holoenzyme. TPX2 is responsible for activating Aurora-A during mitosis, ensuring proper cell division. Disruption of the interface with TPX2 is therefore a potential target for novel anticancer drugs that exploit the increased sensitivity of cancer cells to mitotic stress. Here, we investigate the interface using coprecipitation assays and isothermal titration calorimetry to quantify the energetic contribution of individual residues of TPX2. Residues Tyr8, Tyr10, Phe16, and Trp34 of TPX2 are shown to be crucial for robust complex formation, suggesting that the interaction could be abrogated through blocking any of the three pockets on Aurora-A that complement these residues. Phosphorylation of Aurora-A on Thr288 is also necessary for high-affinity binding, and here we identify arginine residues that communicate the phosphorylation of Thr288 to the TPX2 binding site. With these findings in mind, we conducted a high-throughput X-ray crystallography-based screen of 1255 fragments against Aurora-A and identified 59 hits. Over three-quarters of these hits bound to the pockets described above, both validating our identification of hotspots and demonstrating the druggability of this protein–protein interaction. Our study exemplifies the potential of high-throughput crystallography facilities such as XChem to aid drug discovery. These results will accelerate the development of chemical inhibitors of the Aurora-A/TPX2 interaction.
Metadata
Item Type: | Article |
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Authors/Creators: |
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Copyright, Publisher and Additional Information: | © 2017 American Chemical Society. This document is the Accepted Manuscript version of a Published Work that appeared in final form in ACS Chemical Biology, copyright © American Chemical Society after peer review and technical editing by the publisher. To access the final edited and published work see https://doi.org/10.1021/acschembio.7b00537. |
Keywords: | high throughput crystallography; fragment based screening; protein-protein interaction; protein kinase; XChem |
Dates: |
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Institution: | The University of Leeds |
Funding Information: | Funder Grant number Cancer Research UK C24461/A12772 Cancer Research UK C24461/A23302 |
Depositing User: | Symplectic Publications |
Date Deposited: | 24 Oct 2017 15:51 |
Last Modified: | 18 Oct 2018 00:38 |
Status: | Published |
Publisher: | American Chemical Society |
Identification Number: | 10.1021/acschembio.7b00537 |
Open Archives Initiative ID (OAI ID): | oai:eprints.whiterose.ac.uk:122975 |